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problem in centrifuge of protein

problem in centrifuge of protein - Protein Science

problem in centrifuge of protein - Post Questions and Discuss Protein Structure and Function, Protein Localization etc.. Please do not post Proteomic questions here, there is a Proteomic Forum below.


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Old 09-20-2007, 12:03 AM
Pipette Filler
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Default problem in centrifuge of protein



hi
I`m making 1- centrifuge the meat tissue hemogenate at 1400 rpm
2- then recentrifuge at 15000 rpm for 15 minutes under cooling
3-then put 40 micro of supernatent + 10 micro loading buffer
4-then denaturate at 95 c for 5 minute
5-then centrifuge at 15000 rpm under cooling
every time I make this step
but now I make the step number 2 but not under cooling
then at the end of work the bands of protein is very weak
so I`m asking is not cooling desruct the protein
also I ask can I neglect the step number 5 that mean put the supernatent+loading buffer and denaturate only then inject it in the well
please reply me quickly to repeat the work
thanks
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